INHIBITION OF INOSITOL 1,4,5-TRISPHOSPHATE FORMATION BY CYCLIC-GMP IN CULTURED AORTIC ENDOTHELIAL-CELLS OF THE PIG

1 In cultured endothelial cells of the pig the endothelium-derived relaxing factor (EDRF) releasing agent thrombin (2 u ml-1) caused a significant increase in basal levels of both guanosine 3': 5'-cyclic monophosphate (cyclic GMP) and inositol 1,4,5-trisphosphate (IP3). This increase was time dependent, with peak levels occurring at 2 min and returning towards basal values after 5 min. 2 Pretreatment of the cells with the EDRF inhibitors haemoglobin (1-mu-M) or L-N(G)-nitro arginine (50-mu-M) significantly reduced the cyclic GMP response to thrombin. Both agents also resulted in significant elevations in basal levels of IP3. The IP3 response to thrombin was significantly enhanced at all time points by haemoglobin and at 5 min for L-N(G)-nitro arginine, when compared with the response to thrombin alone. 3 Pretreatment of the cells with either sodium nitroprusside (10-mu-M) or atrial natiuretic peptide (1-mu-M) caused a significant elevation of basal cyclic GMP levels. Although subsequent exposure to thrombin caused a further increase in cyclic GMP, which together with the rise induced by the previous two agents was significantly greater than the increase caused by thrombin alone, the incremental increase induced by thrombin was markedly less in the presence of nitroprusside or atrial natriuretic peptide. Both these agents, as well as 8-bromo cyclic GMP, resulted in a significant suppression of the IP3 response to thrombin. 4 These findings show that one mechanism for the inhibitory effect of cyclic GMP on EDRF release from endothelium may be through the inhibition of IP3 formation in response to EDRF releasing agents.